The GABAA receptor is the target for the inhibitory transmitter GABA, and as such is the primary mediator of inhibition in the CNS. This receptor is a pentamer, composed of varying combinations of alpha (1-6), beta (1-3) and gamma (1-3) or delta subunits. Each specific combination of subunits yields a receptor with distinct pharmacological and biophysical properties, which in turn can influence the excitability of circuits in the CNS areas such as hippocampus, with implications for mood, learning and pathological states such as epilepsy.
The GABAA receptor is also the target for sedative drugs such as benzodiazepine tranquilizers, sedatives, anesthetics and alcohol. In addition, it is the target for a class of steroids which normally act to increase inhibition in the brain. Steroids such as THP (allopregnanolone) are more potent than benzodiazepine tranquilizers in enhancing GABAA receptor function, and as such can effectively decrease anxiety. At high doses, this steroid can also be anesthetic. THP is a metabolite of progesterone, and fluctuates across the ovarian cycle, but is also released during stress.
GABAA Receptor Plasticity
However, GABAAreceptor populations are not static, but change dynamically across periods of prolonged activation in response to neurosteroid fluctuations. This has been shown at puberty, across the ovarian cycle, pregnancy or chronic stress, when alterations in GABAA receptor structure and function occur. In particular, the neurosteroid THP exhibits “withdrawal” properties, when declining levels trigger increased expression of the relatively underexpressed α4 subunit. GABAA receptors containing this subunit have faster kinetics, yielding receptors which produce less total inhibition, but are uniquely responsive to steroids and alcohol.
Figure 2. Section through the CA1 hippocampus showing staining for the α4 subunit, which is increased at puberty in female mice.
Our most recent findings show that steroid fluctuations at puberty increase expression of α4βδ GABAA receptors on the dendrites of CA1 hippocampal pyramidal cells. These receptors uniquely respond to the steroid THP with a different outcome depending on the direction of chloride flux through the receptor. Under conditions of outward chloride current, as found in hippocampus at puberty, THP decreases current through these receptors. (In other areas of the brain, THP increases inward current.) This effect was dependent upon arginine 353, a positively charged amino acid, in the intracellular loop of α4. THP decreased tonic inhibition in the CA1 hippocampus, thereby increasing anxiety at puberty. Because THP is released by stress, prolonged restraint stress also increased anxiety at puberty. This effect was not seen in mice lacking expression of α4βδ GABAA receptors, suggesting that α4βδ GABAA receptors underlie stress-induced mood swings at puberty.
Other Ongoing Studies
Our laboratory is involved in elucidating the functional effects of increases in α4β2δ GABAA receptor expression using electrophysiological, molecular, behavioral and imaging techniques. We have shown that α4βδ GABAA receptors are uniquely responsive to low concentrations of alcohol, and are continuing to characterize this effect. Because α4βδ GABAA receptors are localized to the dendrites of CA1 hippocampal pyramidal cells, we are testing the role of GABA-mediated inhibition in cognition and in vitro models of plasticity, such as LTP in collaboration with Dr. Armin Stelzer in the department. We are also investigating the molecular aspects of steroid actions on α4-containing GABAA receptors using single-site mutagenesis and expression in recombinant systems in collaboration with Dr. Keith Williams in the department. In addition, we have begun to study the regulation of α4 GABAA receptors, using fluorescent imaging techniques to study receptor expression. We are currently collaborating with Dr. Chiye Aoki at NYU, in order to understand the localization pattern of α4 and δ subunits.
Figure 3. Current clamp recordings from a CA1 hippocampal pyramidal cell showing that THP reduces excitability before puberty (top), but increases neuronal excitability at the onset of puberty (lower trace)
Figure 4. The intracellular loops of α1 and α4 GABAA receptor subunits have limited homology (<10%). Mutation of the positively charged amino acid arginine 353 (at arrow) to a neutral glutamine prevented the inhibitory effect of THP on α4β2δ GABAA receptors. (* indicate identical amino acids for both α1 and α4.) α1 sequence: NP899155 [Rattus norvegicus]
– Identical to human α4 sequence: AAB46866 [human, fetal brain]
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Smith, S. S., Gong, Q. H., Li, X., Moran, M. H., Bitran, D., Frye, C. A., and Hsu, F.-C. (1998). Withdrawal from 3α-OH-5α-pregnan-20-one withdrawal using a pseudopregnancy model alters the kinetics of hippocampal GABAA-gated current and increases the GABAA receptor α4 subunit in association with increased anxiety. J. Neurosci. 18, 5275-5284.
Sundstrom-Poromaa, I., Smith, D. H., Gong, Q.-H., Li, X., Light, A., Wiedmann, M., Williams, K., and Smith, S. S. (2002). Hormonally regulated α4β2δ GABAA receptors are a target for alcohol. Nature Neurosci. 5, 721-722.
Hsu, F.-C., and Smith, S. S. (2003). Progesterone withdrawal decreases paired-pulse inhibition in rat hippocampus: An effect dependent upon GABAA receptor α4 subunit upregulation. J. Neurophysiol. 89, 186-198.
Hsu, F.-C., Waldeck, R., Faber, D. S., and Smith, S. S. (2003). Neurosteroid effects on GABAergic synaptic plasticity in hippocampus. J. Neurophysiol. 89, 1929-1940.
Smith, S. S., and Gong, Q. H. (2005). Neurosteroid administration and withdrawal alter GABAA receptor kinetics in CA1 hippocampus of female rats. J. Physiol. 564, 421-436.
Zhou, X., and Smith, S. S. (2007). Steroid requirements for regulation of the a4 subunit of the GABAA receptor in an in vitro model. Neurosci. Lett. 411, 61-66.
Shen, H., Gong, Q.-H., Aoki, C., Yuan, M., Ruderman, Y., Dattilo, M., Williams, K., and Smith, S. S. (2007). Reversal of neurosteroid effects at α4β2δ GABAA receptors triggers anxiety at puberty. Nature Neurosci. 10, 469-477.
Shen, H., Sabaliauskas, N., Sherpa, A., Fenton, A. A., Stelzer, A., Aoki,, C., Smith, S. S. (2010) A critical role for α4β2δ GABAA receptors in shaping learning deficits at puberty in mice. Science 327, 1515-1518.
Qi Hua Gong, M.S.
Aarti Kuver, Ph.D.,
Nicole Sabaliauskas, B.S.,
Hui Shen, M.D., Ph.D.
Research Associate Professor
Ad hoc NTRC NIH study section
Manuscript reviewer for numerous journals